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Atypical Modes of CTCF Binding Facilitate Tissue-Specific and Neuronal Activity-Dependent Gene Expression States

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Abstract

Multivalent binding of CTCF to variable DNA sequences is thought to underlie its ability to mediate diverse cellular functions. CTCF typically binds a 20 base-pair consensus DNA sequence, but the full diversity of CTCF binding sites (CBS) within the genome has not been interrogated. We assessed CTCF occupancy in cultured cortical neurons and observed surprisingly that ~ 22% of CBS lack the consensus CTCF motif. We report here that sequence diversity at most of these atypical CBS involves degeneracy at specific nucleotide positions within the consensus CTCF motif, which likely affect the binding of CTCF zinc fingers 6 and 7. This mode of atypical CTCF binding defines most CBS at gene promoters, as well as CBS that are dynamically altered during neural differentiation and following neuronal stimulation, revealing how atypical CTCF binding could influence gene activity. Dynamic CBS are distributed both within and outside loop anchors and TAD boundaries, suggesting both looping-dependent and independent roles for CTCF. Finally, we describe a second mode of atypical CTCF binding to DNA sequences that are completely unrelated to the consensus CTCF motif, which are enriched within the bodies of tissue-specific genes. These tissue-specific atypical CBS are also enriched in H3K27ac, which marks cis-regulatory elements within chromatin, including enhancers. Overall, these results indicate how atypical CBS could dynamically regulate gene activity patterns during differentiation, development, and in response to environmental cues.

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Data Availability

Original data produced for this manuscript, including raw and processed CTCF and SMC1 ChIP-seq data, are openly available from the NCBI GEO database at https://www.ncbi.nlm.nih.gov/geo/query/acc.cgi?acc=GSE235972, reference number GSE235972.

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Acknowledgements

We would like to thank our colleges in the Madabhushi Lab, Lance Heady, Lahiri Konada, and Ilse Delint-Ramirez, for their helpful feedback on this manuscript.

Funding

This work was supported by NIMH under grant number MH120132 (RM) and CPRIT under grant number RP210041(MC).

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Authors and Affiliations

  1. Department of Psychiatry, University of Texas Southwestern Medical Center, Dallas, TX, USA

    Morgan Crewe, Amir Segev, Richard Rueda & Ram Madabhushi

  2. Department of Neuroscience, University of Texas Southwestern Medical Center, Dallas, TX, USA

    Morgan Crewe, Amir Segev, Richard Rueda & Ram Madabhushi

  3. Department of Cell Biology, University of Texas Southwestern Medical Center, Dallas, TX, USA

    Morgan Crewe, Amir Segev, Richard Rueda & Ram Madabhushi

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All authors contributed to the study conception and design. Material preparation and data collection were performed by Morgan Crewe and Richard Rueda. Data analysis was performed by Morgan Crewe, Amir Segev, and Ram Madabhushi. The first draft of the manuscript was written by Morgan Crewe and Ram Madabhushi, and all authors commented on previous versions of the manuscript. All authors read and approved the final manuscript.

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Correspondence to Ram Madabhushi.

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Crewe, M., Segev, A., Rueda, R. et al. Atypical Modes of CTCF Binding Facilitate Tissue-Specific and Neuronal Activity-Dependent Gene Expression States. Mol Neurobiol 61, 3240–3257 (2024). https://doi.org/10.1007/s12035-023-03762-5

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